OMV-mediated protection to AMPs. Relative survival of WT (solid line) and ΔyieM (dashed line) E.coli after 2 h treatment with the indicated concentrations of polymyxin B (A) and colistin (B). (C) Cultures of mid-log phase WT E. coli were simultaneously treated with the indicated antibiotic (polymyxin B (PMB) 1.5 μg/mL and colistin (COL) 1.0 μg/mL) and either no OMVs (black bars) or with OMVs purified from WT E.coli (4 μg/mL) (grey bars). (D) To titrate OMV-mediated protection, indicated concentrations of WT OMVs were co-incubated in media for 2 h with indicated concentrations of polymyxin B and the media cleared of OMVs by centrifugation. Polymyxin B activity remaining in the media was assessed by adding the pretreated media to a mid log-phase culture of WT E. coli, incubating for 2 h, and plating for CFU. Relative growth (% Survival) was determined by dividing the CFU/mL obtained from antibiotic-treated cultures by the CFU/mL from cultures without antibiotic. (n = 9 for all experiments).